Muscular dystrophies involving the dystrophin–glycoprotein ... · Combined positional cloning and candidate gene approaches have been used to identify an increasing number of genes

349

The dystrophin–glycoprotein complex (DGC) is a multisubunitcomplex that connects the cytoskeleton of a muscle fiber to itssurrounding extracellular matrix. Mutations in the DGC disruptthe complex and lead to muscular dystrophy. There are a fewnaturally occurring animal models of DGC-associatedmuscular dystrophy (e.g. the dystrophin-deficient mdx mouse,dystrophic golden retriever dog, HFMD cat and theδ-sarcoglycan-deficient BIO 14.6 cardiomyopathic hamster)that share common genetic protein abnormalities similar tothose of the human disease. However, the naturally occurringanimal models only partially resemble human disease. Inaddition, no naturally occurring mouse models associated withloss of other DGC components are available. This hasencouraged the generation of genetically engineered mousemodels for DGC-linked muscular dystrophy. Not only haveanalyses of these mice led to a significant improvement in ourunderstanding of the pathogenetic mechanisms for thedevelopment of muscular dystrophy, but they will also beimmensely valuable tools for the development of noveltherapeutic approaches for these incapacitating diseases.

AddressesHoward Hughes Medical Institute, Department of Physiology andBiophysics, Department of Neurology, University of Iowa College ofMedicine, Iowa City, Iowa 52242, USA*e-mail: [email protected]

Current Opinion in Genetics & Development 2002, 12:349–361

0959-437X/02/$ — see front matter© 2002 Elsevier Science Ltd. All rights reserved.

AbbreviationsCMD congenital muscular dystrophyDGC dystrophin–glycoprotein complexDMD/BMD Duchenne or Becker muscular dystrophyFCMD Fukuyama congenital muscular dystrophyHFMD hypertrophic feline muscular dystrophyLGMD limb–girdle muscular dystrophyMdx X-chromosome-linked muscular dystrophyMEB muscle–eye–brain diseasemyd myodystrophynNOS neuronal nitric oxide synthase

IntroductionMuscular dystrophy is a general term that describes agroup of inherited and gradually debilitating myogenic disorders. Genetically, the pattern of inheritance can beX-linked recessive as in Duchenne or Becker musculardystrophy (DMD/BMD), autosomal dominant as inlimb–girdle muscular dystrophy type 1 (LGMD type 1), orautosomal recessive as in limb–girdle muscular dystrophytype 2 (LGMD type 2). DMD is the most common type of muscular dystrophy affecting approximately 1 out of3500 males whereas the limb–girdle muscular dystrophiesaffect roughly 1 out of 20,000. Clinically, the muscular

dystrophies are a heterogeneous group of disorders.Patients with DMD have a childhood onset phenotype anddie by their early twenties as a result of either respiratoryor cardiac failure, whereas patients with BMD have moderate weakness in adulthood and may have normal lifespans. The limb–girdle muscular dystrophies have a highly variable onset and progression, but the unifyingtheme among the limb–girdle muscular dystrophies is theinitial involvement of the shoulder and pelvic girdle muscles. Moreover, muscular dystrophies may or may notbe associated with cardiomyopathy [1–4].

Combined positional cloning and candidate geneapproaches have been used to identify an increasing numberof genes that are mutated in various forms of muscular dystrophy. According to the genetic basis, muscular dystro-phies have now been reclassified and close to 30 geneshave been implicated to cause muscular dystrophy (see [5]for review; see also Table 1). The first gene to be clonedwas the dystrophin gene that is mutated in DMD andBMD [6]. Soon after the discovery of dystrophin, the dystrophin–glycoprotein complex (DGC) was identifiedand these studies opened up a new avenue of musculardystrophy research [7–9]. Within the past couple of years,several targeted mouse models for DGC-associated muscular dystrophy have been generated and these mousemodels, which are the focus of this review, have signifi-cantly contributed to understanding the pathogeneticmechanisms of muscular dystrophy.

Dystrophin–glycoprotein complexThe DGC is a large complex of membrane-associated proteins that is critical for the integrity of skeletal musclefibers. This complex consists of dystrophin, the dystroglycans(α and β), the sarcoglycans (α, β, γ and δ), sarcospan, the syntrophins (α1, β1, β2; γ1- and γ2-syntrophins have beenidentified in neurons) and α-dystrobrevin [7,8,10–21].Dystrophin binds to cytoskeletal actin and to the transmem-brane protein β-dystroglycan; the extracellular domain ofβ-dystroglycan binds to the peripheral membrane protein,α-dystroglycan; and α-dystroglycan binds laminin-2 in thebasal lamina [22–24] (see Figure 1). Furthermore, α-dystro-glycan has been shown to bind the heparane sulfateproteoglycans agrin and perlecan and to the chondroitin sulfate proteoglycan biglycan [25–28]. In addition, α-dystro-glycan was recently shown to bind neurexin in neurons [29].Perlecan and agrin are, along with laminin-2, also present inthe basement membrane of the skeletal muscle. Thus, theDGC serves as a link between the extracellular matrix andthe subsarcolemmal cytoskeleton and the DGC is thought toprotect muscle cells from contraction-induced damage[30–31]. In agreement with this hypothesis, mutations in

Muscular dystrophies involving the dystrophin–glycoprotein complex:an overview of current mouse modelsMadeleine Durbeej and Kevin P Campbell*

genes encoding dystrophin, all four sarcoglycans and thelaminin α2 chain are responsible for DMD/BMD, limb–girdlemuscular dystrophy type 2C-F and congenital muscular dystrophy (CMD), respectively [6,11,14–17,32,33]. Not onlydoes the DGC have structural roles but it may also play a rolein signaling. Several signaling molecules bind DGC corecomponents. Grb2, a signal transduction adapter protein,binds β-dystroglycan [34]. The filamins have been implicatedas signal transducers and a member of the filamin family, fil-amin 2, interacts with γ- and δ-sarcoglycan [35]. Furthermore,dystrophin interacts with two classes of cytoplasmic mole-cules, the syntrophins [36,37] and dystrobrevins, which have

been implicated in signaling [21]. α1-syntrophin contains aPDZ-domain (a protein–protein interaction motif) that inter-acts with at least two sarcolemmal proteins involved in signaltransduction, neuronal nitric oxide synthase (nNOS) [38] andvoltage-gated sodium channels [39,40]. Similarly, the PDZ-domain of β2-syntrophin (expressed at the neuromuscularjunction where it binds the dystrophin homologue utrophin)interacts with the PDZ-domain-containing microtubule-associated serine/threonine kinases MAST205 and SAST[41]. α-dystrobrevin binds syntrophins in addition to dystrophin and is a substrate for tyrosine kinases [42,43].Moreover, biochemical evidence was recently presented for

350 Genetics of disease

Table 1

Muscular dystrophies and corresponding mouse models.

DiseaseMode of inheritance andgene locus Gene product Mouse models

X-linked MDDuchenne/Becker MD XR Xp21 Dystrophin MdxEmery–Dreifuss MD XR Xp28 Emerin –

Limb–girdle MDLGMD 1A AD 5q31 Myotilin –LGMD 1B AD 1q11 Lamin A/C Lmna–/– [121]LGMD 1C AD 3p25 Caveolin-3 Cav3–/–

LGMD 1D AD 6q23 ? –LGMD 1E AD 7q32 ? –LGMD 1F AD 5q31 ? –LGMD 2A AR 15q15 Calpain-3 Capn3–/– [122]LGMD 2B AR 2p13 Dysferlin SJL [123]LGMD 2C AR 13q12 �-sarcoglycan Sgcg–/–

LGMD 2D AR 17q12 �-sarcoglycan Sgca–/–

LGMD 2E AR 4q12 �-sarcoglycan Sgcb–/–

LGMD 2F AR 5q33 �-sarcoglycan Sgcd–/–

LGMD 2G AR 17q11 Telethonin –LGMD 2H AR 9q31 TRIM31 [124] –LGMD 2I AR 19q13 Fukutin-related protein [125] –

Distal MDMiyoshi myopathy AR 2p13 Dysferlin SJL [123]Tibial MD AD 2q31 ? –

Congenital MDClassical or pure CMD AR 6q22 Laminin �2 dyFukuyama CMD AR 9q31 Fukutin –MDC1C�7 integrin CMD

ARAR

19q1312q13

Fukutin-related protein [125]�7 integrin

–Itga7–/–

Ulrich CMDWalker Warburg syndrome [127]Rigid spine CMD

ARARAR

??1p35

Collagen VI �2 [126]?Selenoprotein N [128]

–––

Muscle–eye–brain disease AR 1p32 POMGnT1 –

Other forms of MDEmery-Dreifuss MD AD 1q11 Lamin A/C Lmna–/– [121]Bethlem myopathy AD 21q22 Collagen V1 �1 Col6�1–/– [129]Bethlem myopathy AD 21q22 Collagen V1 �2 –Bethlem myopathy AD 2q37 Collagen V1 �3 –? ? ? Collagen XV [130] Col15�1–/–

EB and MD AR 8q24 Plectin Plectin–/– [131]Facioscapulohumeral MD AD 4q35 ? –Scapuloperoneal MD AD 12q21 ? –Oculopharyngeal MD AD 14q11.2 Poly A binding protein 2 –Myotonic dystrophy AD 19q13 Myotonin-protein kinase/Six5 Six5–/– [132,133]

A summary list of muscular dystrophies, their Mendelian inheritance pattern, chromosomal location, mutated proteins and available mousemodels. MD, muscular dystrophy; LGMD, limb–girdle muscular dystrophy; CMD, congenital muscular dystrophy; EB, epidermolysis bullosa; XR,X-linked recessive; AD, autosomal dominant; AR, autosomal recessive. For primary references on the muscular dystrophies, their mode ofinheritance, gene locus, mutated proteins and available mouse models, please see [5] and main text.

an association of α-dystrobrevin with the sarcoglycan–sarcospancomplex, indicating that the sarcoglycan complex is linked tonNOS signaling via α-syntrophin/α-dystrobrevin [44].

All vertebrates seem to have well-defined sequences encodingdystrophin and its homologues, utrophin and DRP-2.Dystrophin-like sequences have also been identified in inver-tebrates such as amphioxus, sea squirt, starfish, scallop, fruit flyand nematode [45]. In each case, phylogenetic analysesshowed the invertebrate dystrophins to be orthologues of thelast common ancestor of dystrophin, utrophin and DRP-2.Given the diversity of the remainder of the components of thevertebrate DGC, how much simpler is the invertebrate counterpart? The release of the complete sequences of theDrosophila melanogaster and Caenorhabditis elegans genomeshave afforded the opportunity to assess the invertebrate reper-toire of the DGC, using a combination of in vitro and in silicotechniques [46,47] (Table 2). In summary, seventeen knownhuman/mouse components (three dystrophin-related proteins,two dystrobrevins, five sarcoglycans, five syntrophins, one

dystroglycan and one sarcospan) appear to be reduced to eightin Drosophila (one dystrophin, one dystrobrevin, three sarco-glycans, two syntrophins, one dystroglycan and no sarcospan)[46]. Furthermore, C. elegans retains all essential human/mousecounterparts of the DGC, but with less diversity [47].

Among the DGC components, only dystrophin and thesarcoglycans are linked to muscular dystrophy. To date, nohuman mutations have been found in dystroglycan, sarcospan, the syntrophins or the dystrobrevins. Mouse models for each of the core components of the DGC exist,however, and each mouse model (for a summary, see Table 3)will be discussed in relationship to muscular dystrophy.

Animal models for deficiency of dystrophinand dystrophin-binding proteinsDystrophinThe mdx (X-chromosome-linked muscular dystrophy)mouse is the best-characterized mouse model for musculardystrophy: >500 papers have been published in its analysis.

Muscular dystrophies involving the dystrophin–glycoprotein complex Durbeej and Campbell 351

Figure 1

The DGC in skeletal muscle is composed ofdystrophin, the dystroglycans (α, β), thesarcoglycans (α, β, γ, δ), sarcospan, thesyntrophins (α, β1) and dystrobrevin (α).There is a growing number of proteinsreported to be associated with the DGC inthe muscle cell. Depicted are nNOS, whichinteracts with the syntrophin complex andlaminin-2, which is one of many extracellularligands of α-dystroglycan. Several forms ofmuscular dystrophy arise from primarymutations in genes encoding components ofthe DGC. Mutations in dystrophin, all foursarcoglycans and the laminin α2 chain areresponsible for DMD/BMD, LGMD type 2C-Fand CMD, respectively. In addition, severalforms of CMD are caused by abnormalglycosylation of α-dystroglycan (not illustrated).

2C

2D

2E

2FLGMD

Dystroglycancomplex

Dystrobrevin

nNOS

Dystrophin β1

DMD/BMD

Syntrophins

Sarcoglycancomplex

α

α

β

β

δγ

Sarcospan

Laminin-2CMD

α

Current Opinion in Genetics & Development

As a result of a point mutation in exon 23 of the dystrophingene, the mdx mouse is missing dystrophin [48]. Absence ofdystrophin in skeletal muscle also affects the expression ofthe other DGC components at the sarcolemma [49].Although this mouse has proved to be a valuable model forDMD, the progressive muscle-wasting disease presents itselfin a much milder form than in humans [50] at least in certainmuscles. The most affected muscle in the mdx mouse is thediaphragm, which reproduces the degenerative changes ofmuscular dystrophy [51] and the specific twitch force, specifictetanic force and maximum power are all significantlyreduced in diaphragm [52]. The mdx mice show signs ofmuscular dystrophy in other muscles during their first sixweek of life but subsequently show little weakness and havea near-normal lifespan. This partial ‘recovery’ is as a result ofsubstantial regeneration. The mdx mouse adapts to muscledegeneration with an expansion of the satellite cell populationand muscle hypertrophy. Transcription factors seem to beimportant for this successful regeneration. Mdx mice lackingthe muscle-specific transcription factor MyoD show a more

severe dystrophy due to a deficiency in regenerative capabilitiesof the muscle [53]. Likewise, mdx mice deficient in themyocyte nuclear factor, which is selectively expressed insatellite cells, exhibit an exacerbated dystrophic phenotypeas a result of satellite-cell dysfunction [54].

Another possible explanation for the mild phenotype ofthe mdx mouse is that the homologous protein utrophincompensates for the lack of dystrophin. Indeed, mice lacking both dystrophin and utrophin show many signs ofDMD in humans: they have a reduced life-span (dyingbetween 4 and 20 weeks), they suffer from severe muscleweakness with joint contractures, pronounced growthretardation, kyphosis and cardiomyopathy, suggesting thatdystrophin and utrophin play complementary roles [55,56].The dystrophic phenotype of the utrn–/–/mdx mouse wassubsequently ameliorated by skeletal-muscle-specificexpression of truncated utrophin, indicating thatutrn–/–/mdx mice succumb to a skeletal muscle defect andthat their reduced life-span is not as a result of either


Table 2

DGC components in various organisms.

ProteinMus musculusAcc No

DrosophilaAcc No

C. elegansAcc No

Danio rerioAcc No

Dystroglycan DystroglycanX86073

Dystroglycan (DG)AF277390

DystroglycanZ68011

DystroglycanBF157619

�-sarcoglycan �-sarcoglycanAF064081

��-sarcoglycan (SCG��

AF277391��-sarcoglycanAF077544

�


�-sarcoglycan (SGC�)AF277392

�-sarcoglycanAF099925

�-sarcoglycanAA495110



AF277393��-sarcoglycanZ68314

�

�-sarcoglycan �-sarcoglycanAB024923


AF277393��-sarcoglycanZ68314

�-sarcoglycanAI877617



AF277391��-sarcoglycanAF07754

�-sarcoglycanAW281123

Sarcospan SarcospanNM_010656

Not determined ? ?

Dystrophin andhomologues

Dystrophin,Utrophin,DRP-2

Dystrophin (DYS)X99757

Dystrophin (dys-1)AJ012469

DystrophinAJ012469

NM_007868YI2229U43520

Dystrobrevins �-, �-dystrobrevinNM_010087AJ003007

Dystrobrevin (DYB)AF277387

Dystrobrevin (dyb-1)AJ131742

?

Syntrophins �1-, �1-, �2-syntrophins

Syntrophin-1 (SYN1)AF277388

�1-syntrophinZ81072

�1-syntrophinAW280633

NM_009228U89997U40572a

�1-, �1-syntrophinsNM_018967a

NM 018968a

Syntrophin-2 (SYN2)AF277388

a Human sequences. A summary list of DGC sequences from mouse, fruitfly, nematode and zebrafish. Accession (Acc) numbers (underlined)were extracted from GenBank entries for genomic and cDNA clones (see also [46,47]).

cardiac or neurogenic components [57]. Although the mdxmouse has the capability to adapt to muscle degenerationand utrophin to some extent can compensate for the loss ofdystrophin, it should be noted that the serum levels of creatine kinase are elevated in these mice [58], which is atypical diagnostic criteria for muscular dystrophies.

Other than dystrophin, several shorter isoforms are alsogenerated from the dystrophin gene through differentialpromoter usage. Transcripts from four internal dystrophinpromoters produce proteins of 260, 140, 116 and 71 kDa(Dp260, Dp140, Dp116 and Dp71) [59–62]. Dp260 isexpressed predominantly in retina [61]. Dp140 is localizedin brain and kidney [62,63] and Dp116 is expressed exclu-sively in Schwann cells [59]. Dp71 is expressed at highlevels in almost all tissues except skeletal muscle [60]. Yet,mice with a targeted inactivation of Dp71 display no obvious phenotype [64]. The mdx2cv-5cv are other mutantmice generated by chemical mutagenesis using N-ethyl-nitrosurea [65]. The mdx3cv has a mutation at the 3′ end ofthe dystrophin gene, thus resulting in deficiency of theshorter isoforms. Absence of all dystrophin isoforms alongwith utrophin, however, does not worsen the dystrophicphenotype compared to the utrn–/–/mdx mouse [66].

Attempts have also been made to produce a dystrophingene knockout mouse (mdx52). Araki et al. deleted exon 52of dystrophin to produce a mouse with a similar phenotypeto the mdx mouse [67].

Syntrophinα1-syntrophin is strongly expressed at the sarcolemma andis a core protein of the DGC [7,37]. Yet, mice deficient inα1-syntrophin display no gross histological changes in theskeletal muscle and muscle contractile properties are notaltered in these mice [68,69]. nNOS is misplaced from thesarcolemma of these mice and so is aquaporin-4, a mercurial-insensitive water-selective channel [70]. Aquaporin-4 isalso absent in skeletal muscle of mdx mice [71]. Yet, therole of aquaporin-4 in the pathogenesis of muscular dystrophyis unclear as aquaporin-4-deficient mice maintain normalmuscle function [72]. α1-syntrophin is also expressedabundantly at the neuromuscular junction, and recentstudies reveal that absence of α1-syntrophin leads to structurally aberrant neuromuscular synapses deficient inutrophin [69].

DystrobrevinThe best-studied functions of the DGC involve structuralstabilization of the sarcolemma: mutations of several DGC components appear to cause muscular dystrophy by disassembling the complex and compromising the linkagebetween the extracellular matrix of the fibers to itscytoskeleton. Mice deficient in α-dystrobrevin maintainthe expression of the DGC at the sarcolemma; yet, thesemice develop a mild muscular dystrophy. Analysis ofα-dystrobrevin mice revealed that muscular dystrophymight also develop as a result of impaired DGC-dependentsignaling. The mechanism behind the disrupted signaling


Table 3

Summary of DGC-associated mouse models.

Genotype (protein absent) Life-span Skeletal dystrophy Cardiomyopathy

Sgca–/– (�-sarcoglycan) >1 yr Moderate NoneSgcb–/– (�-sarcoglycan) >1yr Severe SevereSgcg-/- (�-sarcoglycan) 20 wks Severe SevereSgcd–/– (�-sarcoglycan) >1yr Severe SevereSspn–/– (sarcospan) >1yr None NoneDG–/– (dystroglycan) Embryonic lethal NA NADG chimeric (dystroglycan absent in skeletal and cardiac muscle) >1yr Moderate? SevereMyd (large) Reduced Moderate NoneMdx (dystrophin) >1yr Mild/moderate MildMdx2cv (dystrophin, Dp260) >1yr Mild/moderate MildMdx3cv (dystrophin, Dp71, Dp116, Dp140, Dp260) >1yr Mild/moderate MildMdx4cv (dystrophin, Dp140, Dp260) >1yr Mild/moderate MildMdx5cv (dystrophin) >1yr Mild/moderate MildMdx52 (dystrophin, Dp140, Dp260) >1 yr Mild/moderate NoneDp71–/– >6 months None NoneMnf/mdx (myocyte nuclear factor, Dystrophin) ~3 wks Severe NTMyoD/mdx (MyoD, dystrophin) 1 yr Severe SevereNNOS–/– (neuronal nitric oxide synthase) >1yr None NonenNOS/mdx >1yr Mild/moderate MildUtrn–/– (utrophin) >1yr None NoneUtrn–/–/mdx (utrophin, dystrophin) 4–20 wks Severe SevereUtrn–/–/mdx3cv (utrophin, dystrophin, Dp 260, Dp140, Dp116, Dp71) 4–20 wks Severe SevereAdbn–/– (�-dystrobrevin) >1 yr Mild MildAdbn–/–/mdx (�-dystrobrevin, dystrophin) 8–10 months Moderate ModerateUtrn–/–/mdx/adbn–/– (utrophin, dystrophin, �-dystrobrevin) 3–11 wks Severe Severe�1-syntrophin–/– >1yr None NoneCav-3–/– (caveolin-3) >30 wks Mild None

NA, not applicable; NT, not tested. (For references, please see main text.)

remains to be determined but nNOS may be involved.During exercise, nNOS activity is stimulated to producecyclic GMP, which is necessary to maintain the blood flowin the active muscle. In electrically stimulated muscle ofα-dystrobrevin deficient mice, cyclic GMP levels are notaffected in agreement with the finding that nNOS is displaced from the muscle membrane [21]. Yet, loss ofnNOS alone is unlikely to account for the dystrophy inα-dystrobrevin-deficient mice, as mice lacking nNOS arenot dystrophic [73]. Moreover, the genetic loss of nNOSdoes not alter the pathogenesis of mdx mice [74]. However,recent studies suggest that at least part of the muscledegeneration observed in DMD patients may result fromthe reduced production of muscle-membrane-associatednNOS. This reduction may lead to improper control of thevasculature and eventual local muscle ischemia [75].

Caveolin-3Caveolin-3 is a muscle-specific protein integrated in thecaveolae, which are small invaginations of the plasmamembrane. Mutations in the human caveolin-3 gene causemild muscular dystrophy (LGMD 1C) [76]. Moreover,caveolin-3 has by immunoprecipitation experiments beenshown to interact with dystrophin [77] but it is not an integral component of the DGC [78]. Very recently, caveolin-3-deficient mice were generated [79,80]. Thesemice exhibit very mild myopathic changes [79,80].Interestingly, Lisanti and co-workers showed that cave-olin-3 expression is required for correct targeting of theDGC to cholesterol-sphingolipid raft domains/caveolae innormal muscle fibers [80].

Animal models for dystroglycan deficiencyDystroglycan was originally isolated from skeletal musclebut has since been shown to be expressed in a wide varietyof tissues and is now considered to be the most broadlyexpressed DGC component [10,81]. Besides muscle, dystroglycan is expressed at high levels in developing andadult tissues, typically in cell types that adjoin basementmembranes such as epithelial and neural tissue [82–84].Early in vitro work demonstrated a role for dystroglycan inepithelial morphogenesis [82]. In 1997, the dystroglycangene was disrupted in mouse [85]. Dystroglycan nullembryos fail to progress beyond the early egg cylinderstage of development and are characterized by structuraland functional perturbations of Reichert’s membrane, oneof the earliest basement membranes that form in therodent embryo. Subsequent work demonstrated a role fordystroglycan in the formation of the basement membraneof the embryoid body [84]. Because dystroglycan nullembryos die early in development prior to gastrulation (i.e. long before any muscle has formed) it is not possibleto analyze the consequences of dystroglycan deficiency inmuscle. To overcome this, Carbonetto and co-workers generated chimeric mice, lacking dystroglycan in skeletalmuscle [86]. Interestingly, these mice develop progressivemuscle pathology with changes emblematic to musculardystrophies in humans. In addition, many neuromuscularjunctions are disrupted in these mice. Thus, dystroglycanis necessary for myofiber stability and synapse differentiationor stability. Surprisingly, the basement membrane in thechimeric mice is not grossly perturbed. Yet, the sarcoglycansand dystrophin are absent from the dystroglycan-deficient


Figure 2

Microfil perfusion of the vasculature.(a,b) Vessels of 4-week-old heart anddiaphragm from wild-type mice show smoothlytapered vessel walls. (c,d) In contrast, vesselsof the heart and diaphragm from Sgcb-nullmice (deficient in β-sarcoglycan) shownumerous constrictions (denoted by arrows).

Wt heart Wt diaphragm

Sgcb null heart Sgcb null diaphragm

(a) (b)

(c) (d)


muscle fibers suggesting that that the sarcolemmal basementmembrane interactions are weakened and/or disorganized.

It is becoming increasingly clear that posttranslational modifi-cations of muscle cell proteins, in particular α-dystroglycan,are important for normal muscle function. Grewal et al. [87••]recently presented very intriguing data on the importance ofcorrect glycosylation of α-dystroglycan. The myodystrophymouse (myd) harbors a mutation in the glycosyltransferase,Large, which leads to altered glyco-sylation of α-dystroglycan(however, with no apparent shift in the molecular weight of α-dystroglycan) and mutant mice develop a progressivemyopathy [87••]. Furthermore, muscle–eye–brain disease(MEB) is a type of congenital muscular dystrophy associatedwith loss-of-function mutations in the gene encoding a glycosyltransferase, POMGnT1 [88••]. POMGnT1 is a glycosylation enzyme that participates in the synthesis ofO-mannosyl glycan, a modification that is rare in mammalsbut is present in α-dystroglycan. Kano et al. [89•] recentlyreported a deficiency of α-dystroglycan, but not β-dystro-glycan in MEB patients. A similar selective secondary deficiency of heavily glycosylated α-dystroglycan was alsonoted in Fukuyama congenital muscular dystrophy (FCMD;in which the gene encoding fukutin is affected) [90]. In summary, a growing body of evidence indicates that somemuscular dystrophies may be caused by abnormal glyco-sylation of α-dystroglycan. The mechanistic basis for thesedisorders is yet to be determined.

Animal models for laminin-2 deficiencyIn skeletal muscle, α-dystroglycan binds to the basementmembrane protein laminin-2 (composed of laminin α2, β1 andγ1 chains) [24]. About 50% of the patients diagnosed with classical CMD show a primary deficiency of the lamininα2 chain and basement membrane perturbations [91]. Severalmouse models for laminin-2 deficiency now exist, includingthe dy (dystrophia-muscularis) mouse, originally identified atthe Jackson Laboratory [24,92,93], and an allelic mutant of thedy mouse, dy2J [94]. Neither of these mouse models exhibitsa complete deficiency of laminin α2 chain. Yet, they both display a muscular dystrophy, although the muscular dystrophyin the dy2J presents itself in a milder form compared to the dymouse. Several laboratories have also generated null mutantsfor laminin α2 chain (dy3K, dyW) and all of these mouse modelspresent a severe muscular dystrophy caused by the failure toform a laminin scaffold, which is necessary for basement membrane structure and for interactions with the DGC andintegrins [95,96]. Interestingly, a mini agrin gene, whichretained high-affinity binding sites for the laminins that areupregulated in dyW mice (laminin α4 and laminin α5 chains)and α-dystroglycan compensated for the loss of laminin α2chain [97••]. This suggests that even a non-homologous high-affinity link between dystroglycan and the extracellular matrixis sufficient to prevent muscular dystrophy.

Animal models for integrin deficiencyLaminins also bind integrins, which are a large family of heterodimeric transmembrane cell surface receptors that

function in a wide variety of cell interactions [98]. In skeletalmuscle, the α7β1 integrin is the predominant integrin thatbinds laminin-2 [99]. It is possible that the functions of theDGC and the integrin α7 complex in skeletal muscle tosome extent overlap. Mice lacking integrin α7 display a mildmyopathy [100] and, interestingly, mice lacking both integrin α7 and dystrophin develop a severe dystrophy anddie between 3–4 weeks of age (U Mayer, unpublished data).The fact that the expression of integrin α7 transcript andprotein is increased in mdx mice and in DMD patients further suggests that the integrin α7β1 may compensate for theabsence of the DGC [101,102]. Indeed, transgenic expressionof the α7β1 integrin reduces muscular dystrophy andrestores viability in the dystrophic utrn–/–/mdx mouse [103].


Figure 3

Schematic diagram of sarcoglycan complexes in muscle. (a) Mouseskeletal muscle was stained with β-sarcoglycan antibodies. (b) Normal skeletal muscle contains two sarcoglycan complexes: anα-sarcoglycan and an ε-sarcoglycan containing complex. The former iscomposed of α-, β-, γ- and δ-sarcoglycan whereas the latter iscomposed of at least ε-, β- and δ-sarcoglycan and perhapsγ-sarcoglycan (denoted by a broken line). Note that only thesarcoglycan complexes (and not the entire DGC) are illustrated.(c) Mouse cardiac muscle was stained with β-sarcoglycan antibodies.(d) Two sarcoglycan complexes are also present within cardiacmuscle. (e) Pulmonary artery is positively stained with β-sarcoglycanantibodies. (f) The smooth muscle sarcoglycan complex is composedof ε-, β-, γ- and δ-sarcoglycan.

αβγδ εβγδ

α β

γ δ

ε β

γ δ

αβγδ εβγδ

β

δ

ε β

γ δ

εβγδ

ε β

γ δ

α

γ

Skeletal muscle(a) (b)

(d)

(f)Smooth muscle(e)

Cardiac muscle(c)


Although α7β1 integrin is the predominant integrin thatbinds laminin-2 in skeletal muscle, other integrins areimportant for normal muscle function too. Mice chimericfor the expression of integrin α5 (receptor for fibronectin)also develop a myopathy [104].

In summary, these data suggest that both the DGC andintegrins are involved in anchoring the muscle fiber to itsextracellular matrix and that disruption of this anchorageresults in muscle instability and subsequently cell death.

Animal models for sarcoglycan–sarcospandeficiencyThe sarcoglycan complex is a group of single-pass transmembrane proteins (α-, β-, γ- and δ-sarcoglycan) thatis tightly associated with sarcospan to form a subcomplexwithin the DGC [105]. Although the exact function of thesarcoglycan–sarcospan complex is not known, it is wellestablished that mutations in any of α-, β-, γ- and δ-sarco-glycan genes result in distinct forms of muscular dystrophynow collectively called sarcoglycanopathies [11,14–18,32].A primary mutation in any one of these genes may lead toeither total or partial loss of that sarcoglycan as well as asecondary deficiency of the other sarcoglycans [106].Furthermore, sarcospan expression is dependent on properexpression of the sarcoglycans [58,105,107]. To date, however, no human mutations have been found in the sarcospan gene, nor have any unclassified muscular dystro-phies been mapped to the chromosomal location ofsarcospan [107]. Moreover, sarcospan-deficient mice main-tain their sarcolemmal expression of DGC and maintainnormal muscle function [108].

The sarcoglycans are primarily expressed in muscle. α-, β-,γ-, and δ-sarcoglycan are, along with sarcospan, expressedin skeletal and cardiac muscle [109]. In smooth muscle, onthe other hand, a unique sarcoglycan–sarcospan complex is expressed. The smooth muscle sarcoglycan complex iscomposed of ε-sarcoglycan (an α-sarcoglycan homologue)instead of α-sarcoglycan, along with β-sarcoglycan, γ-sarcoglycan and δ-sarcoglycan [81,109,110•]. The sarco-glycan–sarcospan complex is in smooth muscle alsoassociated with dystroglycan. However, dystroglycan insmooth muscle is differentially glycosylated compared todystroglycan in skeletal muscle [109].

Recent data from several laboratories, including ours, havedemonstrated that proper expression of the sarcoglycans inskeletal muscle is a necessity for normal skeletal musclefunction [58,111–114,115••]. Furthermore, we have recentlydemonstrated that proper expression of the sarcoglycans insmooth muscle is also of great importance for normal skeletaland cardiac muscle function [112,115••].

Mice deficient in α-sarcoglycan (Sgca null mice), which isexclusively expressed in striated muscle, develop a progressive muscular dystrophy and a concomitant deficiency in β-, γ- and δ-sarcoglycan along with sarcospan

in skeletal muscle. Furthermore, α-dystroglycan is greatlydestabilized at the sarcolemma, indicating that membraneexpression of the sarcoglycans is a prerequisite for membrane targeting and stabilization of α-dystroglycan.The sarcoglycan–sarcospan complex is also significantlyreduced in cardiac muscle of Sgca null mice. Yet, thesemice do not develop cardiomyopathy [58]. The reason forthis became clear when β- and δ-sarcoglycan-deficientmice were generated and analyzed in our laboratory[112,115••]. Mice deficient in β- and δ-sarcoglycan (Sgcbnull mice and Sgcd null mice, respectively) that areexpressed in both striated and smooth muscle also developa progressive muscular dystrophy, which seems to be moresevere than in Sgca null mice. Large, focal areas of necrosis/fibrosis are present in skeletal muscle of Sgcb and Sgcdnull mice, a phenomenon that is not detected in Sgca null mice. Both Sgcb and Sgcd null mice also develop cardiomyopathy and morphologically this is recognized by extensive areas of necrosis/fibrosis in the hearts.Additionally, the serum levels for cardiac-specific troponin I are elevated in these mice in agreement withthe presence of acute myocardial necrosis that precedesthe formation of fibrotic lesions [112,115••,116••].Furthermore, deficiency of β- and δ-sarcoglycan leads toperturbed expression of the entire sarcoglycan–sarcospanand dystroglycan complexes not only in striated musclebut also in vascular smooth muscle. The loss of the sarco-glycan–sarcospan complex in smooth muscle and thepresence of the focal areas of necrosis in skeletal and car-diac muscle prompted us to analyze the blood vessels moreclosely in Sgcb and Sgcd null mice. Using the Microfil per-fusion technique, we found vascular irregularities in theform of arterial constrictions in both heart and skeletalmuscle and also in the kidneys of Sgcb and Sgcd null mice([112,115••]; M Durbeej, KP Campbell, unpublished data)(see Figure 2). Importantly, the disturbance of the vasculatureprecedes the onset of ischemic-like lesions. Moreover, novascular perturbations are present in Sgca null mice.α-sarcoglycan is not expressed in smooth muscle — thesmooth muscle sarcoglycan is thus intact in Sgca null mice.Still, Sgca null mice are dystrophic. Together, these datasuggest that muscle degeneration does not cause the vascular phenotype. Instead, loss of the sarcoglycan–sar-cospan complex in smooth muscle of blood vessels resultsin vascular irregularities that aggravate skeletal pathologyand initiate heart pathology as a result of diminished delivery of oxygen and nutrients [112,113••,116••].

Biochemical analysis of sarcoglycan deficient micerevealed the presence of a new sarcoglycan complex in skeletal and cardiac muscle ([114,115••]; M Durbeej,KP Campbell, unpublished data). This additional sarcoglycancomplex is composed of at least ε-, β-, and δ-sarcoglycan,but neither dystrophin or utrophin [115••] (Figure 3).Thus, β- and δ-sarcoglycan mutations, but not α-sarcoglycanmutations, affect the expression of this complex, which issubsequently absent in Sgcb and Sgcd null mice but notSgca null mice. Hence, loss of the ε-sarcoglycan complex


may also contribute to the more severe pathology seen inSgcb and Sgcd null mice. Whether γ-sarcoglycan is part ofthe ε-sarcoglycan complex is not clear. ε-sarcoglycanexpression remains in γ-sarcoglycan-deficient mice [111].However, immunoprecipitation experiments indicate that γ-sarcoglycan is indeed part of the ε-sarcoglycan complex [114]. Clearly, more studies are needed to clarifythis discrepancy.

In summary, a complex pathogenetic mechanism for thedevelopment of LGMD 2E and F can be postulated.Deficiency of β- and δ-sarcoglycan causes loss of the sarco-glycan–sarcospan and dystroglycan complexes as well as lossof the ε-sarcoglycan-containing complex in striated muscle.Since the linkage between the extracellular matrix and themuscle cell is perturbed, the skeletal and cardiac musclemembranes become unstable. Thus, the muscle cells may bemore prone to ischemic damage that develops as a conse-quence of the absent smooth muscle sarcoglycan complex.

Although α-, β-, γ- and δ-sarcoglycans are part of the samecomplex, the individual sarcoglycans may exhibit differentfunctional roles. Several lines of evidence support thisidea. First, ecto-ATPase activity has been noted forα-sarcoglycan, indicating that α-sarcoglycan might functionin controlling the ATP concentration at the surface of themuscle cell [117]. Second, mice deficient for γ-sarcoglycanexhibit a severe muscular dystrophy and cardiomyopathy[111]. Without γ-sarcoglycan, β- and δ-sarcoglycan areunstable at the muscle membrane and α-sarcoglycan isseverely reduced. The expression of dystroglycan, however,is not altered and thus the link between the extracellularmatrix and the intracellular cytoskeleton appears unaffected.Moreover, γ-sarcoglycan-deficient muscle show normalresistance to mechanical strain, normal peak isometric andtetanic force generation and no evidence for contraction-induced injury after exercise [118]. Thus, a nonmechanicalmechanism (perhaps signaling) may be responsible forγ-sarcoglycan-deficient muscular dystrophy. These resultsare in sharp contrast to muscles of Sgca, Sgcb and Sgcd nullmice, which show abnormal resistance to stretch, and adecrease in specific force generation ([58,112]; M Durbeej,KP Campbell, unpublished data).

ConclusionsAnimal models lacking each component of the DGC haveprovided many new insights into the development of muscular dystrophy. Specifically, we have learned thatmuscular dystrophy can develop when DGC core compo-nents such as sarcoglycans, dystroglycan and dystrobrevinsare missing from the skeletal muscle. In addition, musculardystrophy can also develop when laminin-2 and integrinsare absent. More importantly, analysis of these mousemodels has unraveled novel pathogenetic mechanisms forthe development of muscular dystrophy. For example, perturbation of vascular function together with the disruptionof the ε-sarcoglycan complex contribute to the increasedseverity of mouse models of LGMD type 2E and F.

Furthermore, these animal models will be tremendouslyvaluable tools for testing novel therapeutic approaches. Infact, the feasibility of sarcoglycan gene transfer for sarco-glycanopathies has already been investigated using someof the sarcoglycan-deficient mice. These studies demon-strate that mutations in individual sarcoglycan componentscan be corrected in vivo ([119•,120•]; M Durbeej,KP Campbell, unpublished data). Moreover, the mousemodels will be of immense benefit for evaluating possibledrug therapies for muscular dystrophy and cardiomyopathy.For example, nicorandil, a vascular smooth muscle relaxant,prevents the acute onset of myocardial necrosis in Sgcdnull mice [112]. Furthermore, recent studies in our laboratorysuggest that verapamil, a pharmacological agent withvasodilator properties, successfully prevents cardiomyopathyin sarcoglycan-deficient mice by abolishing the vasculardysfunction [116••]. Taken together, the successful treat-ment of cardiomyopathy using agents with clinicalrelevance in mouse models lacking the smooth musclesarcoglycan–sarcospan complex connotes that effortstoward drug therapies can be of tremendous benefit preventing certain forms of hereditary cardiomyopathy andperhaps muscular dystrophy.

Acknowledgements We would like to thank all members of the Campbell laboratory for theircritical reading of the manuscript and for fruitful discussions. MuscularDystrophy Association supported this work. KP Campbell is an Investigatorof the Howard Hughes Medical Institute.

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88. Yoshida A, Kobayashi K, Manya H, Taniguchi K, Kano H, Mizuno M,•• Inazu T, Mitsuhashi H, Takahashi S, Takeuchi M et al.: Muscular

dystrophy and neuronal migration disorder caused by mutationsin a glycosyltransferase, POMGnT1. Dev Cell 2001, 1:717-724.

An important paper elucidating the genetic basis for MEB. Loss-of-functionmutations are described in the glycosyltransferase POMGnT1, an enzymethat participates in O-mannosyl glycan synthesis. Mammalian O-mannosylglycosylation is a rare type of post-translational modification but interestinglyit occurs on α-dystroglycan. Together with [87••], these data indicate thataltered glycosylation of α-dystroglycan is one cause of muscular dystro-phies. See also [89•].

89. Kano H, Kobayashi K, Herrmann R, Tachikawa M, Manya H, Nishino I,• Nonaka I, Straub V, Talim B, Voit T et al.: Deficiency of

αα-dystroglycan in muscle-eye-brain disease. Biochem Biophys Res Comm 2002, 291:1283-1286.

A follow up study to [88••]. Immunohistochemical and immunoblot analysesdemonstrate that α-dystroglycan, but not β-dystroglycan is absent in skeletalmuscle of MEB patients.

90. Hayashi YK, Ogawa M, Tagawa K, Noguchi S, Ishihara T, Nonaka I,Arahata K: Selective deficiency of alpha-dystroglycan inFukuyama-type congenital muscular dystrophy. Neurology 2001,10:115-121.

91. Tomé FM: The Peter Emil Becker Award lecture: The saga ofcongenital muscular dystrophy. Neuropediatrics 1998, 30:55-65.

92. Michelson AM, Russell E, Harman PJ: Dystrophia muscularis:a hereditary primary myopathy in the house mouse. Proc AcadNatl Sci USA 1955, 41:1079-1084.

93. Xu H, Christmas P, Wu X, Wewer U, Engvall, E: Defective musclebasement membrane and lack of M-laminin in the dy/dy mouse.Proc Natl Acad Sci USA 1994, 91:5572-5576.

94. Xu H, Wu R, Wewer U, Engvall E: Murine muscular dystrophycaused by a mutation in the laminin alpha 2 (Lama2) gene. NatGenet 1994, 8:297-302.

95. Miyagoe Y, Hanaoka K, Nonaka I, Hayasaka M, Nabeshima Y,Arahata K, Nabeshima Y, Takeda S: Laminin alpha2 chain-nullmutant mice by targeted disruption of the Lam2 gene: a newmodel of merosin (laminin 2)-deficient congenital musculardystrophy. FEBS Lett 1997, 415:33-39.

96. Kuang W, Xu H, Vachon PH, Liu L, Loechel F, Wewer UM, Engvall E:Merosin-deficient congenital muscular dystrophy. Partial geneticcorrection in two mouse models. J Clin Invest 1998, 102:844-852.

97. Moll J, Barzaghi P, Lin S, Bezakova G, Lochmuller H, Engvall E,•• Muller U, Ruegg MA: An agrin minigene rescues dystrophic

symptoms in a mouse model for congenital muscular dystrophy.Nature 2001, 413:302-307.

An intriguing approach to alleviate muscle pathology in the laminin-α2-deficient dyw mouse. An agrin minigene was designed to restore the linkbetween dystroglycan and the extracellular matrix. Transgenic overexpres-sion of the minigene in the skeletal muscles of dyw mice prevented musclewasting. These data provide evidence that a non-homologous protein incombination with a rational protein design could replace the function of amissing protein.

98. Hynes RO: Integrins: versatility, modulation, and signaling in celladhesion. Cell 1992, 69:11-25.

99. Burkin DJ, Kaufman SJ: The αα7ββ1 integrin in muscle developmentand disease. Cell Tiss Res 1999, 296:183-190.

100. Mayer U, Saher G, Fassler R, Bornemann A, Echtermeyer F, von derMark H, Miosge N, Poschl E, von der Mark K: Absence of integrin αα7causes a novel form of muscular dystrophy. Nat Genet 1997,17:318-323.

101. Hodges BL, Hayashi YK, Nonaka I, Wang W, Arahata K, Kaufman SJ:Altered expression of the αα7ββ1 integrin in human and murinemuscular dystrophies. J Cell Sci 1997, 110:2873-2881.

102. Cohn RD, Mayer U, Saher G, Herrmann R, van der Flier A,Sonnenberg A, Sorokin L, Voit T: Secondary reduction of alpha7Bintegrin in laminin αα2 deficient congenital muscular dystrophysupports an additional transmembrane link in skeletal muscle.J Neurol Sci 1999, 163:140-152.

103. Burkin DJ, Wallace GQ, Nicol KJ, Kaufman DJ, Kaufman SJ:Enhanced expression of the alpha 7 beta 1 integrin reducesmuscular dystrophy and restores viability in dystrophic mice.J Cell Biol 2001, 152:1207-1218.

104. Taverna D, Disatnik MH, Rayburn H, Bronson RT, Yang J, Rando TA,Hynes RO: Dystrophic muscle in mice chimeric for expression ofαα5 integrin. J Cell Biol 1998, 143:849-859.

105. Crosbie RH, Lebakken CS, Holt KH, Venzke DP, Straub V, Lee JC,Grady RM, Chamberlain JS, Sanes JR, Campbell KP: Membranetargeting and stabilization of sarcospan is mediated by thesarcoglycan subcomplex. J Cell Biol 1999, 145:153-165.

106. Barresi R, Confalonieri V, Lanfossi M, Di Blasi C, Torchiana E,Mantegazza R, Jarre L, Nardocci N, Boffi P, Tezzon F et al.:Concomitant deficiency of ββ- and γγ-sarcoglycans in 20αα-sarcoglycan (adhalin)-deficient patients: immunohistochemicalanalysis and clinical aspects. Acta Neuropathol 1997, 94:28-35.

107. Crosbie RH, Im LE, Moore SA, Hirano M, Hays AP, Maybaum SW,Collin H, Dovico SA, Stolle CA, Fardeau M et al.: Molecular andgenetic characterization of sarcospan: insights intosarcoglycan–sarcospan domains. Hum Mol Genet 2000,13:2019-2023.

108. Lebakken CS, Venzke DP, Hrstka RF, Consolino CM, Faulkner JA,Williamson RA, Campbell KP: Sarcospan-deficient mice maintainnormal muscle function. Mol Cell Biol 2000, 20:1669-1677.

109. Straub V, Ettinger AJ, Durbeej M, Venzke DP, Cutshall S, Sanes JR,Campbell KP: εε-sarcoglycan replaces αα-sarcoglycan in smoothmuscle to form a unique dystrophin-glycoprotein complex. J BiolChem 1999, 274:27989-27996.

110. Barresi R, Moore SA, Stolle CA, Mendell JR, Campbell KP:• Expression of γγ-sarcoglycan in smooth muscle and its interaction

with the smooth muscle sarcoglycan-sarcospan complex. J BiolChem 2000, 275:38554-38560.

The expression pattern of γ-sarcoglycan in smooth muscle is biochemicallyand genetically evaluated in this paper as it had been unclear whetherγ-sarcoglycan is expressed in smooth muscle. The authors provide convinc-ing evidence that γ-sarcoglycan is a component of the smooth muscle sarcoglycan complex.

111. Hack AA, Ly CT, Jiang F, Clendenin CJ, Sigrist KS, Wollmann RL,McNally EM: γγ-sarcoglycan deficiency leads to muscle membranedefects and apoptosis independent of dystrophin. J Cell Biol1998, 142:1279-1287.

112. Coral-Vasquez R, Cohn RD, Moore SA, Hill JA, Weiss RM,Davisson RL, Straub V, Barresi R, Bansal D, Hrstka RF et al.:Disruption of the sarcoglycan-sarcospan complex in vascularsmooth muscle: a novel mechanism for cardiomyopathy andmuscular dystrophy. Cell 1999, 98:465-474.

113. Araishi K, Sasaoka T, Imamura M, Noguchi S, Hama H,Wakabayashi E, Yoshida M, Hori T, Ozawa E: Loss of thesarcoglycan complex and sarcospan leads to muscular dystrophyin beta-sarcoglycan deficient mice. Hum Mol Genet 1999,8:1589-1598.

114. Liu AL, Engvall E: Sarcoglycan isoforms in skeletal muscle. J BiolChem 1999, 272:38171-38176.

115. Durbeej M, Cohn RD, Hrstka RF, Moore SA, Allamand V, •• Davidson BL, Williamson RA, Campbell KP: Disruption of the

ββ-sarcoglycan gene reveals pathogenetic complexity of limb-girdle muscular dystrophy type 2E. Mol Cell 2000, 5:141-151.

Together with [112], the findings suggest a novel pathogenetic mechanismwhereby disruption of the sarcoglycan–sarcospan complex in vascularsmooth muscle causes disturbance of the vasculature, which in turn leads tothe development of a severe cardiomyopathy and exacerbation of musculardystrophy in β- and δ-sarcoglycan deficient mice. The presence of a distinctε-sarcoglycan complex in skeletal muscle is also presented.

116. Cohn R, Durbeej M, Moore SA, Coral-Vazquez R, Prouty S, •• Campbell KP: Prevention of cardiomyopathy in mouse models

lacking the smooth muscle sarcoglycan-sarcospan complex.J Clin Invest 2001, 107:R1-R9.

An interesting study showing that pharmacological intervention using oraltreatment with verapamil is able to prevent cardiomyopathy in mice with disruption of the smooth muscle sarcoglycan complex.

117. Betto R, Senter L, Ceoldo S, Tarricone E, Biral D, Salviati G:Ecto-ATPase activity of αα-sarcoglycan (adhalin). J Biol Chem1999, 274:7907-7912.

118. Hack AA, Cordier L, Shoturma DI, Lam MY, Sweeney HL,McNally EM: Muscle degeneration without mechanical injury insarcoglycan deficiency. Proc Natl Acad Sci USA 1999,96:10723-10728.


119. Allamand V, Donahue KM, Straub V, Davisson RL, Davidson BL,• Campbell KP: Early adenovirus-mediated gene transfer effectively

prevents muscular dystrophy in alpha-sarcoglycan deficient mice.Gene Therapy 2000, 7:1385-1391.

This paper shows that early adenovirus-mediated gene transfer effectivelyprevents muscular dystrophy in α-sarcoglycan deficient mice. One singleinjection of a first generation adenovirus into the skeletal muscle of neonatemice led to sustained expression of α-sarcoglycan at the sarcolemma oftransduced fibers for at least 7 months.

120. Cordier L, Hack AA, Scott MO, Barton-Davis ER, Gao G, Wilson JM,• McNally EM, Sweeney HL: Rescue of skeletal muscles of

γγ-sarcoglycan-deficient mice with adeno-associated virus-mediated gene transfer. Mol Therapy 2000, 1:119-129.

The authors demonstrate that injections of a recombinant adeno-associatedvirus — in which human γ-sarcoglycan is driven by a muscle creatine kinasepromoter — into skeletal muscle of γ-sarcoglycan-deficient mice result in anoverall improvement of the histological pattern of dystrophy. However, theauthors underscore the need for intervention early in the time course of thedisease process.

121. Sullivan T, Escalante-Alcalde D, Bhatt H, Anver M, Bhat N,Nagashima K, Stewart CL, Burke B: Loss of A-type laminexpression compromises nuclear envelope integrity leading tomuscular dystrophy. J Cell Biol 1999, 147:913-920.

122. Richard I, Roudaut C, Marchand S, Baghdiguian S, Herasse M,Stockholm D, Ono Y, Suel L, Bourg N, Sorimachi H et al.: Loss ofcalpain 3 proteolytic activity leads to muscular dystrophy and toapoptosis-associated IκκBαα/nuclear factor κκB pathwayperturbation in mice. J Cell Biol 2000, 151:1583-1590.

123. Bittner RE, Anderson LV, Burkhardt E, Bashir R, Vafiadaki E,Ivanova S, Raffelsberger T, Maerk I, Hoger H, Jung M et al.: Dysferlindeletion in SJL mice (SJL-Dysf) defines a natural model for limbgirdle muscular dystrophy 2B. Nat Genet 1999, 23:141-142.

124. Frosk P, Weiler T, Nylen E, Sudha T, Greenberg CR, Morgan K,Fujiwara TM, Wrogemann K: Limb-girdle muscular dystrophytype 2H associated with mutation in TRIM32, a putative E3-ubiquitin-ligase gene. Am J Hum Genet 2002, 70:663-672.

125. Brockington M, Yuva Y, Prandini P, Brown SC, Torelli S, Benson MA,Herrmann R, Anderson LV, Bashir R, Burgunder JM et al.: Mutations

in the fukutin-related protein identify limb-girdle musculardystrophy 2I as a milder allelic variant of congenital musculardystrophy MDC1C. Hum Mol Genet 2001, 10:2851-2859.

126. Camacho Vanegas O, Bertini E, Zhang RZ, Petrini S, Minosse C,Sabatelli P, Giusti B, Chu ML, Pepe G: Ullrich scleroatonicmuscular dystrophy is caused by recessive mutations in collagentypeVI. Proc Natl Acad Sci USA 2001, 98:7516-7521.

127. Cormand B, Pihko H, Bayes M, Valanne L, Santavuori P, Talim B,Gershoni-Baruch R, Ahmad A, van Bokhoven H, Brunner HG et al.:Clinical and genetic distinction between Walker-Warburgsyndrome and muscle-eye-brain disease. Neurology 2001,56:1059-1069.

128. Moghadaszadeh B, Petit N, Jaillard C, Brockington M, Roy SQ,Merlini L, Romero N, Estournet B, Desguerre I, Chaigne D et al.:Mutations in SEPN1 cause congenital muscular dystrophy withspinal rigidity and restrictive respiratory syndrome. Nat Genet2001, 29:17-18.

129. Bonaldo P, Braghetta P, Zanetti M, Piccolo S, Volpin D, Bressan GM:Collagen VI deficiency induces early onset myopathy in themouse: an animal model for Bethlem myopathy. Hum Mol Genet1998, 7:2135-2140.

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Muscular dystrophies involving the dystrophin–glycoprotein ... · Combined positional cloning and candidate gene approaches have been used to identify an increasing number of genes